Multiplexed protein quantification with barcoded hydrogel microparticles.
Multiplexed protein quantification with barcoded hydrogel microparticles.
复制标题
DOI：
10.1021/ac1022343
复制
发表时间：
2011-01-01
影响因子：
7.4
通讯作者：
Doyle, Patrick S.
中科院分区：
文献类型：
--
作者：
Appleyard, David C.;Chapin, Stephen C.;Doyle, Patrick S.
我们演示了在复杂介质中使用图形编码的水凝胶微粒对临床相关蛋白板进行敏感和高通量多路检测。结合已建立的抗体捕获技术与微流控合成和分析的进步，我们在单次和多次检测中检测到1 - 8 pg/mL的三种细胞因子（白细胞介素-2，白细胞介素-4和肿瘤坏死因子α），而无需过滤或阻断剂。研究了一系列水凝胶孔隙度，以确保目标和试剂快速扩散到颗粒中，并在冲洗过程和高速微流控扫描过程中保持颗粒的结构完整性。使用异双功能聚乙二醇连接剂将捕获抗体共价结合，仅使用少量有价值的试剂即可一步合成和功能化颗粒。除了使用三种不同类型的单探针颗粒外，利用停止流动光刻（SFL）方法的灵活性，在单个编码颗粒上对上述目标集的三种探针进行空间隔离，从而证明了单颗粒诊断面板的可行性。本研究建立了凝胶-颗粒平台作为有效定量蛋白质靶点的通用工具，并显著推进了将水凝胶底物和颗粒阵列的优势扩展到临床蛋白质组学领域的努力。
We demonstrate the use of graphically encoded hydrogel microparticles for the sensitive and high-throughput multiplexed detection of clinically relevant protein panels in complex media. Combining established antibody capture techniques with advances in both microfluidic synthesis and analysis, we detected 1−8 pg/mL amounts of three cytokines (interleuken-2, interleuken-4, and tumor necrosis factor alpha) in single and multiplexed assays without the need for filtration or blocking agents. A range of hydrogel porosities was investigated to ensure rapid diffusion of targets and reagents into the particle as well as to maintain the structural integrity of particles during rinsing procedures and high-velocity microfluidic scanning. Covalent incorporation of capture antibodies using a heterobifunctional poly(ethylene glycol) linker enabled one-step synthesis and functionalization of particles using only small amounts of valuable reagents. In addition to the use of three separate types of single-probe particles, the flexibility of the stop-flow lithography (SFL) method was leveraged to spatially segregate the three probes for the aforementioned target set on an individual encoded particle, thereby demonstrating the feasibility of single-particle diagnostic panels. This study establishes the gel−particle platform as a versatile tool for the efficient quantification of protein targets and significantly advances efforts to extend the advantages of both hydrogel substrates and particle-based arrays to the field of clinical proteomics.
登录
查看更多内容
影响因子：
7.4
作者：
Broder, Graham R.;Ranasinghe, Rohan T.;Morgan, Hywel
通讯作者：
Morgan, Hywel
影响因子：
30.5
作者：
Sokol, Caroline L.;Barton, Gregory M.;Medzhitov, Ruslan
通讯作者：
Medzhitov, Ruslan
影响因子：
4.4
作者：
Yates, John;Rovis, Flavia;Garden, O. A.
通讯作者：
Garden, O. A.
影响因子：
2.8
作者：
Lee, Woojin;Choi, Dongkil;Koh, Won-Gun
通讯作者：
Koh, Won-Gun
影响因子：
12.3
作者：
Peck, David;D Crawford, Emily;Ross, Kenneth N.;Stegmaier, Kimberly;Golub, Todd R.;Lamb, Justin
通讯作者：
Lamb, Justin